稳定干扰长链非编码RNA LINC01224结直肠癌细胞株的建立及其对细胞凋亡的影响
Acta Universitatis Medicinalis Anhui（2022）
Abstract
目的 建立稳定干扰长链非编码RNA(lncRNA)LINC01224表达的结直肠癌LoVo和SW620细胞株,并探讨下调LINC01224表达对结直肠癌细胞凋亡的影响.方法 使用GEPIA2数据库分析LINC01224在结直肠癌组织中的表达情况;qPCR法检测LINC01224在10种人结直肠癌细胞中的表达水平.3种不同的LINC01224 siRNA分别转染人结直肠癌LoVo细胞,取抑制LINC01224表达效果最显著的siRNA序列构建LINC01224 shRNA慢病毒载体.在HEK293T细胞内包装成重组慢病毒颗粒,再感染LoVo和SW620细胞,经嘌呤霉素筛选后以有限稀释法获得稳定干扰LINC01224的单克隆细胞.MTS法检测细胞增殖能力,流式细胞术检测细胞凋亡率.结果 LINC01224在结直肠癌组织中的表达高于正常结直肠组织,其在10种结直肠癌细胞中的表达也高于正常结直肠上皮细胞HCOEPic.siRNA-3对LoVo细胞内LINC01224表达的抑制率高于siRNA-1和siRNA-2.故选择siRNA-3设计LINC01224 shRNA.与对照组(sh-NC组)相比,稳定干扰LINC01224组(sh-LINC01224组)LoVo和SW620细胞内LINC01224的表达水平降低(P<0.01),其细胞生长速度减慢(P<0.01),凋亡率也增加(P<0.01).结论 成功构建LINC01224的shRNA慢病毒干扰载体,该载体能够稳定感染LoVo和SW620细胞,下调LINC01224表达并诱导细胞凋亡.
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